lonotropic glutamate receptors control a wide variety of normal neuronal processes, including learning and memory. In addition, activation of these important neurotransmitter receptors is involved in a number of neurodegenerative diseases, notably stroke and epilepsy. Thus, drugs targeted toward glutamate receptors would be of considerable therapeutic value. Analysis of the transmembrane topology led to the realization that each subunit is made up of a series of modules, and the module that binds glutamate can be produced in bacteria as a soluble protein (S1S2 domain). The S1S2 domain binds agonists and antagonists with approximately the same affinity as the intact receptor and serves as an excellent system for studying the binding domain. The crystal structure of the S1S2 domain of the GluR2 subunit has been solved in the presence of several agonists and antagonists by E. Gouaux and collaborators, which provided a major breakthrough in the molecular understanding of these receptors. The purpose of this proposal is to complement the known three-dimensional structure with dynamic information from NMR spectroscopy. Preliminary studies have shown that the S1S2 domain of GluR2 is well suited to these studies. The backbone resonances have been assigned, and preliminary results with glutamate in the binding site provide a picture of the portions of the binding site that are mobile, and a potential mechanism for glutamate dissociation. Further studies of the backbone and sidechain dynamics, in the presence of various agonists and an antagonist, and at different temperatures, will provide a detailed molecular picture of the energetics of ligand binding, and the dynamics associated with binding and, potentially, of the transduction of binding to channel function. A second goal is to measure the conformational change upon binding in solution. As shown by the crystal structures, the S1S2 domain is a bilobe structure that closes upon ligand binding. However, the degree of lobe closure has been shown in similar systems to be influenced by crystal packing and lobe dynamics, suggesting that lobe closure in solution may differ from that in the crystal. Residual dipolar coupling measurements will be used to study the orientation of the two lobes, with different ligands in the binding site and at different temperatures. This should allow an extrapolation of the three-dimensional structure to the solution state and provide additional information on the energetics of binding. The results of these studies will shed light on the binding site of an important glutamate receptor subunit, and provide essential information for further drug development.